Synthesis of 2-amino-5-substituted-1,3,4-oxadiazoles through the electrochemical oxidation of semicarbazone was carried out at platinum anode at room temperature under controlled potential electrolysis in an undivided cell assembly. The electrolysis were carried out in the non aqueous medium acetic acid and lithium perchlorate was used as a supporting electrolyte. The antimicrobial activities of the compounds have been studied by screening the compounds against two gram negative organisms Klebsiella pneumoniae, Escherichia coli, and two gram positive organisms Bacillus subtilis and Streptococcus aureus and antifungal activity against Aspergillus niger and Crysosporium pannical and results have been compared with the standard antibacterial Streptomycin and antifungal Griseofulvin.

1,3,4-oxadiazole is a versatile lead molecule for designing potential bioactive agents. The 1,3,4-oxadiazole derivatives have been found to exhibit diverse biological activities such as hypotensive1 anti-microbial2-4, anti-HIV2-4, anti-fungal5-6 anti-inflammatory7 antimitotic activity8 and muscle relaxant9

It have been found in the líterature study that the methods for synthesis of oxadiazole 1 include bromine oxidation of semicarbazide derivative and the cyclodesulfurization of acylthiosemicarbazide derivatives in the solution using I2/NaOH or 1,3-dicyclohexylcarbodimide (DCC)10-13 as well as mercury(II) acetate (Hg(OAc)2) or yellow mercury(II) oxide HgO14-16 Evans17 synthesized oxadiazole derivatives by rapid parallel synthesis in efficient one-pot preparation using resin-bound reagents. All these methods are usually carried out in various different synthetic steps and require the heating at higher temperature. The handling of these reagents is not only difficult but also very hazardous to environment. The each stage of the reaction including extraction and purification of the producís from the mixture requires great precautions.

These reports including our earlier work18-23 prompted us to synthesize 2-amino-5-substituted-1,3,4-oxadiazole derivatives through the electrochemical oxidation of semicarbazone 4 in the hope of getting potent biodynamic agents and evaluate their antimicrobial activity.

EXPERIMENTAL

Analysis and measurements

The reagents used for the preparation of oxadiazoles were of Merck producís. Spectroscopic grade solvents were used for spectral measurements. The carbón, hydrogen and nitrogen contents in each sample were performed at RSIC, CDRI Lucknow. The IR spectra were recorded on a Perkin-Elmer 783 spectrophotometer KBr pellets. 1H NMR and 13C NMR spectra were recorded on a Bruker DRX 300 (300MHz) FT spectrometer in CDCl3 using TMS as internal reference. Chemical shifts are reported in ppm downfield from TMS as internal reference. Elemental analyses were carried out in the Elementar VarioEL III instrument. Melting points were determined on a VEB Wagetechink Rapio PHMK05 instrument and are uncorrected. Water used was doubly distilled.

General procedure for the synthesis of compounds (1a-1n) Semicarbazide hydrochloride 3 (9.0 mmol) and sodium acetate (12.2 mmol) was dissolved in water (10 mL) and then aldehyde 2 (4.16 mmol) was added with continuous stirring. The mixture was left overnight, which gives semicarbazone 4. Now semicarbazone 4 (12.25 mmol), lithium perchlorate (1.3 mmol) and acetic acid (200 mL) was taken in the reaction cell assembly with platinum píate as working as well as counter electrode and saturated calomel electrode as reference electrode. Controlled potential electrolysis24-31 were performed at their corresponding oxidation potentials at room temperature, tabulated as Table 1. The product was extracted from the acetic acid solution to chloroform by the simple solvent extraction technique. 4-6 Fmol-1 of electricity was passed for the electrolysis, which is very small in comparison to energy used in other conventional methods.

All the title compounds were tested for their antimicrobial activity by adopting the experimental method of Benson32. Whatman No. 1 filter paper dises of 6 mm diameter, placed in a Petri dish, were autoclaved. The test compounds in measured quantities (1.0 mg, 0.5 mg) were dissolved in 5 mL of dimethylformamide to produce 200 ppm and 100 ppm solutions, respectively. The filter paper dises were allowed to dry and the amount of the substance per disc was taken as 500 and 250 µg. The bacterial (24 h) and fungal (48 h) cultures from the slants were diluted with sterile water and mixed thoroughly to prepare a clear homogeneous suspensión. These suspensions were uniformly spread on solidified agar (nutrient and potato dextrose agar) medium. The filter paper dises prepared from dimethylformamide medium were carefully placed over the spreaded cultures and incubated at 37 °C for 24 h for bacteria and at 28-30 °C for 48 h for fungi. Paper dises treated with dimethylformamide alone served as control. After the incubation period the plates were examined for inhibition zones. The diameters of inhibition zones (including the diameter of the disc) were measured. All determinations were made in triplícate for each of the compounds and the average valué was taken.

RESULTS AND DISCUSSION

The organic synthesis involving electrochemical techniques under suitable solvents and electrolytes are the basic requirements. The solution phase methods, while successful, were deemed not readily amenable to high throughout synthesis, and thus did not meet our needs. Considering the importance of 1,3,4-oxadiazoles and methods of preparation, 2-amino-5-substituted-1,3,4-oxadiazoles were synthesized by electrooxidative eyelization of semicarbazone as an environmentally benign synthetic method in which aforementioned troublesome were removed and minimized. The electroorganic synthesis of 1,3,4-oxadiazole derivatives 1 is electrochemical eyelization by electrooxidation of semicarbazone 4. The electrochemical oxidation has various merits. The electrooxidation does not require oxidizing reagents and can be performed at room temperature. Electricity provides chemical processes with special attributes, such as enhanced reaction rate, higher yield of puré producís, better selectivity and several ecofriendly advantages.

This electrochemical eyelization gives the oxadiazoles (Scheme 1). Acetic acid was used as a solvent and lithium perchlorate as an electrolyte.

Novel 2-amino-5-substituted-1,3,4-oxadiazoles have been synthesized in excellent yields using the synthetic route outlined in Scheme 1. IR, 1H NMR, 13C NMR and mass spectral data are in agreement with the proposed structures of all synthesized compounds. Lack of 1H NMR resonances observed with NH and NH2 functions in the 1H NMR spectrum of 1a-n proved that ring closure starting from 4 resulted in the formation of 2-amino-1,3,4-oxadiazole ring. This was further substantiated by the 13C NMR data of 1 which showed the peaks at 8 170-173 and 145-150 due to C2 and C5 of oxadiazole respectively. The IR spectrum shows 1600-1620 ern1 for (C=N-N=C) and 1063-1073 cm-1 for (C-O-C) in the compounds 1a-1n which confirmed the synthesis of 1,3,4-oxadiazoles.

Antibacterial Activity

The antibacterial activity of compounds 1a-1n was studied against the growth of Klebsiella pneumoniae, Escherichia coli, (Gram-negative) and Bacillus subtilis, Streptococus aureus (Gram-positive) organisms at the two concentrations (25 and 50 ppm) taking Streptomycin as the standard (Table 2). The majority of the compounds exhibited significant (good) antibacterial activity against E. coli, K. pneumonia, B. subtilis and S. aureus as compared to Streptomycin. The screening results of antibacterial activity revealed that compounds 1e and 1j exhibited approximately similar activity to the standard Streptomycin. Compounds 1f, 1g, 1h, 1i and 1l exhibited slightly less antibacterial activity. Remaining compounds exhibited weak antibacterial activity against all bacterial strains used for our evaluation.

Antifungal Activity

The compounds 1a-1n were screened for their antifungal activity against Aspergillus niger and C. pannical species along with the standard fungicide Griseofulvins 1a-1n (Table 3). The disc diffusion method was followed for screening the compounds atthree concentrations (10, 100 and 1000 ppm). The screening results revealed that all the compounds displayed good antifungal activity against A. niger and C. pannical. However, compounds 1e and 1j showed equal antifungal activity when compared with the Griseofulvins.

The antimicrobial activity of the compounds varied upon the type and position of the substituents at 5-substituted-2-amino-1,3,4-oxadiazole moiety. It can be concluded from the antimicrobial screening results that when 5-substituted-2-amino-1,3,4-oxadiazoles were substituted with aryl halide the antimicrobial activity was altered to an appreciable extent.

CONCLUSION

In conclusión we have developed a convenient method for the synthesis of novel 2-amino-5-substituted-1,3,4-oxadiazole derivatives in excellent yields which is not so easy to achieve by chemical methods. These compounds exhibited significant activity against the growth of bacterial and fungal strain. The antimicrobial activity of the compounds varied upon the type and position of the substituents at 5-substituted-2-amino-1,3,4-oxadiazole moiety. It is evident from the antimicrobial screening results that when 5-substituted-2-amino-1,3,4-oxadiazoles were substituted with aryl halide the antimicrobial activity was altered to an appreciable extent.

ACKNOWLEDGEMENTS

The authorthanks to Sophisticated Analytical Instrument Facility (SAIF) a división of CDRI (Central Drug Research Institute) Lucknow for recording spectra and microanalyses.